Sequence specific complexation of b DNA at sites containing g,c base pairs

A series of eight related analogs of distamycin A has been synthesized. Footprinting and affinity cleaving reveal that only two of the analogs, pyridine-2- car box amide-netropsin (2-Py N) and 1-methylimidazole-2-carboxamide-netrops in (2-ImN), bind to DNA with a specificity different from that of the parent compound. A new class of sites, represented by a TGACT sequence, is a strong site for 2-PyN binding, and the major recognition site for 2-ImN on DNA. Both compounds recognize the G•C bp specifically, although A's and T's in the site may be interchanged without penalty. Additional A•T bp outside the binding site increase the binding affinity. The compounds bind in the minor groove of the DNA sequence, but protect both grooves from dimethylsulfate. The binding evidence suggests that 2-PyN or 2-ImN binding induces a DNA conformational change.

In order to understand this sequence specific complexation better, the Ackers quantitative footprinting method for measuring individual site affinity constants has been extended to small molecules. MPE•Fe(II) cleavage reactions over a 10^5 range of free ligand concentrations are analyzed by gel electrophoresis. The decrease in cleavage is calculated by densitometry of a gel autoradiogram. The apparent fraction of DNA bound is then calculated from the amount of cleavage protection. The data is fitted to a theoretical curve using non-linear least squares techniques. Affinity constants at four individual sites are determined simultaneously. The distamycin A analog binds solely at A•T rich sites. Affinities range from 10^(6)- 10^(7)M^(-1) The data for parent compound D fit closely to a monomeric binding curve. 2-PyN binds both A•T sites and the TGTCA site with an apparent affinity constant of 10^(5) M^(-1). 2-ImN binds A•T sites with affinities less than 5 x 10^(4) M^(-1). The affinity of 2-ImN for the TGTCA site does not change significantly from the 2-PyN value. At the TGTCA site, the experimental data fit a dimeric binding curve better than a monomeric curve. Both 2-PyN and 2-ImN have substantially lower DNA affinities than closely related compounds.

In order to probe the requirements of this new binding site, fourteen other derivatives have been synthesized and tested. All compounds that recognize the TGTCA site have a heterocyclic aromatic nitrogen ortho to the N or C-terminal amide of the netropsin subunit. Specificity is strongly affected by the overall length of the small molecule. Only compounds that consist of at least three aromatic rings linked by amides exhibit TGTCA site binding. Specificity is only weakly altered by substitution on the pyridine ring, which correlates best with steric factors. A model is proposed for TGTCA site binding that has as its key feature hydrogen bonding to both G's by the small molecule. The specificity is determined by the sequence dependence of the distance between G's.

One derivative of 2-PyN exhibits pH dependent sequence specificity. At low pH, 4-dimethylaminopyridine-2-carboxamide-netropsin binds tightly to A•T sites. At high pH, 4-Me_(2)NPyN binds most tightly to the TGTCA site. In aqueous solution, this compound protonates at the pyridine nitrogen at pH 6. Thus presence of the protonated form correlates with A•T specificity.

The binding site of a class of eukaryotic transcriptional activators typified by yeast protein GCN4 and the mammalian oncogene Jun contains a strong 2-ImN binding site. Specificity requirements for the protein and small molecule are similar. GCN4 and 2-lmN bind simultaneously to the same binding site. GCN4 alters the cleavage pattern of 2-ImN-EDTA derivative at only one of its binding sites. The details of the interaction suggest that GCN4 alters the conformation of an AAAAAAA sequence adjacent to its binding site. The presence of a yeast counterpart to Jun partially blocks 2-lmN binding. The differences do not appear to be caused by direct interactions between 2-lmN and the proteins, but by induced conformational changes in the DNA protein complex. It is likely that the observed differences in complexation are involved in the varying sequence specificity of these proteins.

Transition metal clusters supported by multinucleating ligand frameworks as models of biological active sites

This dissertation describes efforts to model biological active sites with small molecule clusters. The approach used took advantage of a multinucleating ligand to control the structure and nuclearity of the product complexes, allowing the study of many different homo- and heterometallic clusters. Chapter 2 describes the synthesis of the multinucleating hexapyridyl trialkoxy ligand used throughout this thesis and the synthesis of trinuclear first row transition metal complexes supported by this framework, with an emphasis on tricopper systems as models of biological multicopper oxidases. The magnetic susceptibility of these complexes were studied, and a linear relation was found between the Cu-O(alkoxide)-Cu angles and the antiferromagnetic coupling between copper centers. The triiron(II) and trizinc(II) complexes of the ligand were also isolated and structurally characterized.

Chapter 3 describes the synthesis of a series of heterometallic tetranuclear manganese dioxido complexes with various incorporated apical redox-inactive metal cations (M = Na⁺, Ca²⁺, Sr²⁺, Zn²⁺, Y³⁺). Chapter 4 presents the synthesis of heterometallic trimanganese(IV) tetraoxido complexes structurally related to the CaMn₃ subsite of the oxygen-evolving complex (OEC) of Photosystem II. The reduction potentials of these complexes were studied, and it was found that each isostructural series displays a linear correlation between the reduction potentials and the Lewis acidities of the incorporated redox-inactive metals. The slopes of the plotted lines for both the dioxido and tetraoxido clusters are the same, suggesting a more general relationship between the electrochemical potentials of heterometallic manganese oxido clusters and their “spectator” cations. Additionally, these studies suggest that Ca²⁺ plays a role in modulating the redox potential of the OEC for water oxidation.

Chapter 5 presents studies of the effects of the redox-inactive metals on the reactivities of the heterometallic manganese complexes discussed in Chapters 3 and 4. Oxygen atom transfer from the clusters to phosphines is studied; although the reactivity is kinetically controlled in the tetraoxido clusters, the dioxido clusters with more Lewis acidic metal ions (Y³⁺ vs. Ca²⁺) appear to be more reactive. Investigations of hydrogen atom transfer and electron transfer rates are also discussed.

Appendix A describes the synthesis, and metallation reactions of a new dinucleating bis(N-heterocyclic carbene)ligand framework. Dicopper(I) and dicobalt(II) complexes of this ligand were prepared and structurally characterized. A dinickel(I) dichloride complex was synthesized, reduced, and found to activate carbon dioxide. Appendix B describes preliminary efforts to desymmetrize the manganese oxido clusters via functionalization of the basal multinucleating ligand used in the preceding sections of this dissertation. Finally, Appendix C presents some partially characterized side products and unexpected structures that were isolated throughout the course of these studies.

Reaction of glucose catalyzed by framework and extraframework tin sites in zeolite beta

The isomerization of glucose into fructose is a large-scale reaction for the production of high-fructose corn syrup, and is now being considered as an intermediate step in the possible route of biomass conversion into fuels and chemicals. Recently, it has been shown that a hydrophobic, large pore, silica molecular sieve having the zeolite beta structure and containing framework Sn⁴⁺ (Sn-Beta) is able to isomerize glucose into fructose in aqueous media. Here, I have investigated how this catalyst converts glucose to fructose and show that it is analogous to that achieved with metalloenzymes. Specifically, glucose partitions into the molecular sieve in the pyranose form, ring opens to the acyclic form in the presence of the Lewis acid center (framework Sn⁴⁺), isomerizes into the acyclic form of fructose and finally ring closes to yield the furanose product. Akin to the metalloenzyme, the isomerization step proceeds by intramolecular hydride transfer from C2 to C1. Extraframework tin oxides located within hydrophobic channels of the molecular sieve that exclude liquid water can also isomerize glucose to fructose in aqueous media, but do so through a base-catalyzed proton abstraction mechanism. Extraframework tin oxide particles located at the external surface of the molecular sieve crystals or on amorphous silica supports are not active in aqueous media but are able to perform the isomerization in methanol by a base-catalyzed proton abstraction mechanism. Post-synthetic exchange of Na⁺ with Sn-Beta alters the glucose reaction pathway from the 1,2 intramolecular hydrogen shift (isomerization) to produce fructose towards the 1,2 intramolecular carbon shift (epimerization) that forms mannose. Na⁺ remains exchanged onto silanol groups during reaction in methanol solvent, leading to a near complete shift in selectivity towards glucose epimerization to mannose. In contrast, decationation occurs during reaction in aqueous solutions and gradually increases the reaction selectivity to isomerization at the expense of epimerization. Decationation and concomitant changes in selectivity can be eliminated by addition of NaCl to the aqueous reaction solution. Thus, framework tin sites with a proximal silanol group are the active sites for the 1, 2 intramolecular hydride shift in the isomerization of glucose to fructose, while these sites with Na-exchanged silanol group are the active sites for the 1, 2 intramolecular carbon shift in epimerization of glucose to mannose.

Towards a Participatory Integrated Assessment Approach for Planning and Managing Natura 2000 Network Sites

36 p.

The role of sites of special scientific interest (SSSIs) in the conservation of British rivers

This paper discusses the particular contribution of the SSSI (Sites of Special Scientific Interest) as a way of nature conservation for rivers. In 1989, the Nature Conservancy Council proposed a dual selection system for selection of rivers; either (1) "Whole river" SSSIs representing the main types of river, or rivers which show classic and representative transitions down their lengths, or (2) "Sectional" SSSIs which are shorter stretches of river with high nature conservation interest. The NCC has recently classified all SSSIs with a river interest into 4 categories: - river SSSIs, river valley SSSIs, river adds interest - where the river clearly adds biological interest to the site, and rivers of incidental interest. The overall length of river SSSIs amounts to almost 1000 km.

Comunidades de anfíbios anuros em duas fitofisionomias do Parque Estadual da Serra do Tabuleiro, estado de Santa Catarina

O estado de Santa Catarina possuía sua área inteiramente coberta por Mata Atlântica, um dos biomas mais ameaçados mundialmente. Mesmo com o crescente aumento de informações sobre os anfíbios no estado, ainda existe muitas lacunas de informações para este grupo e o desconhecimento é alto mesmo no interior das Unidades de Conservação. O Parque Estadual da Serra do Tabuleiro é a maior unidade de conservação de proteção integral de Santa Catarina e ainda assim possui inúmeras ameaças e pressões antrópicas. Os objetivos deste estudo foram estudar os anfíbios anuros em duas fitofisionomias, restinga e floresta ombrófila densa submontana, do Parque Estadual da Serra do Tabuleiro, verificar as diferenças dos parâmetros da comunidade entre três mesohábitats na restinga, compreender em que extensão algumas variáveis estruturais do hábitat afetam a riqueza e abundância dos anuros na restinga e na floresta ombrófila densa submontana, conhecer os anuros de folhiço na área de floresta e, adicionalmente, verificar o atual conhecimento sobre os anuros de folhiço no mundo. Os anuros no parque foram amostrados por meio dos métodos de procura ativa em transecções, armadilhas de queda com cerca guia e parcelas no chão da floresta durante três anos consecutivos com amostragens trimestrais, de julho de 2007 a abril de 2010. As variáves estruturais foram medidas nos locais amostrados enquanto que os dados climáticos foram obtidos da estação meteorológica mais próxima da área de estudo. A revisão sobre os anuros de folhiço no mundo foi realizada por uma busca na base de dados ISI Web if Science entre os anos de 1945 e 2008. Foram registradas 39 espécies de anuros no Parque, sendo 15 espécies para a restinga, 31 espécies na floresta. No folhiço do chão da floresta, a comunidade foi composta por 13 espécies de anuros. No total, 66% dos anuros registrados aqui, foram endêmicos da Mata Atlântica, 17% possuíram distribuição restrita à Santa Catarina e quatro espécies foram consideradas como vulneráveis na lista de espécies ameaçadas de extinção de Santa Catarina. Na restinga, a área aberta e a mata de restinga foram as mais dissimilares com relação à composição e abundância de anuros e também para as variáveis estruturais do hábitat. Na floresta ombrófila densa, a cobertura de dossel e o número de corpos dágua foram as variáveis mais importantes para a riqueza e abundância das espécies de anuros. Também para os anuros de folhiço do parque, a cobertura de dossel foi importante, conjuntamente com a umidade do ar e a profundidade de folhiço. O atual conhecimento sobre os anuros de folhiço no mundo se concentra nas regiões tropicais, sendo que o Brasil possui o maior número de estudos. Algumas tendências para as comunidades de anuros de folhiço puderam ser identificadas, mas estudos adicionais são necessários para que mais inferências possam ser feitas. O presente estudo contribuiu para preencher parte das lacunas de conhecimento existentes para a biodiversidade dos anuros do estado de Santa Catarina e em especial para o Parque Estadual da Serra do Tabuleiro.

Comment: Food web interactions and biomanipulation in Australian reservoirs

The authors have been studying the control of phytoplankton biomass in several Australian reservoirs. To manipulate fish communities in order to reduce phytoplankton biomass, one needs a thorough understanding of processes in the plankton-associated food webs. In contrast to the situation found in lakes of the northern hemisphere, the planktivorous fish of Australian reservoirs are relatively small and they may deplete small rather than large zooplankters, so that animals as large as the adults of Daphnia carinata may avoid predation. This would result in promotion of Daphnia, particularly if fish eliminate its smaller competitors. The aim of biomanipulation should be the establishment and maintenance of a proper ratio of planktivores/piscivores, adequate for water quality requirements. Successful selection of the appropriate ratio for a given reservoir will depend on the extent of our understanding of its food web interactions. For practical application of biomanipulation in management, further development of the food web theory under Australian conditions is needed.

I. Spectroscopic observation of discrete sites in simple liquids. II. Infrared intensity perturbations of hydrogen halide fundamentals in liquid xenon. III. Rotation of HCl in solid rare gases.

Part I

The spectrum of dissolved mercury atoms in simple liquids has been shown to be capable of revealing information concerning local structures in these liquids.

Part II

Infrared intensity perturbations in simple solutions have been shown to involve more detailed interaction than just dielectric polarization. No correlation has been found between frequency shifts and intensity enhancements.

Part III

Evidence for perturbed rotation of HCl in rare gas matrices has been found. The magnitude of the barrier to rotation is concluded to be of order of 30 cm^(-1).

Computational Predictions of G Protein-Coupled Receptor Structures and Binding Sites

G protein-coupled receptors (GPCRs) are the largest family of proteins within the human genome. They consist of seven transmembrane (TM) helices, with a N-terminal region of varying length and structure on the extracellular side, and a C-terminus on the intracellular side. GPCRs are involved in transmitting extracellular signals to cells, and as such are crucial drug targets. Designing pharmaceuticals to target GPCRs is greatly aided by full-atom structural information of the proteins. In particular, the TM region of GPCRs is where small molecule ligands (much more bioavailable than peptide ligands) typically bind to the receptors. In recent years nearly thirty distinct GPCR TM regions have been crystallized. However, there are more than 1,000 GPCRs, leaving the vast majority of GPCRs with limited structural information. Additionally, GPCRs are known to exist in a myriad of conformational states in the body, rendering the static x-ray crystal structures an incomplete reflection of GPCR structures. In order to obtain an ensemble of GPCR structures, we have developed the GEnSeMBLE procedure to rapidly sample a large number of variations of GPCR helix rotations and tilts. The lowest energy GEnSeMBLE structures are then docked to small molecule ligands and optimized. The GPCR family consists of five subfamilies with little to no sequence homology between them: class A, B1, B2, C, and Frizzled/Taste2. Almost all of the GPCR crystal structures have been of class A GPCRs, and much is known about their conserved interactions and binding sites. In this work we particularly focus on class B1 GPCRs, and aim to understand that family’s interactions and binding sites both to small molecules and their native peptide ligands. Specifically, we predict the full atom structure and peptide binding site of the glucagon-like peptide receptor and the TM region and small molecule binding sites for eight other class B1 GPCRs: CALRL, CRFR1, GIPR, GLR, PACR, PTH1R, VIPR1, and VIPR2. Our class B1 work reveals multiple conserved interactions across the B1 subfamily as well as a consistent small molecule binding site centrally located in the TM bundle. Both the interactions and the binding sites are distinct from those seen in the more well-characterized class A GPCRs, and as such our work provides a strong starting point for drug design targeting class B1 proteins. We also predict the full structure of CXCR4 bound to a small molecule, a class A GPCR that was not closely related to any of the class A GPCRs at the time of the work.

Responsabilidade civil por danos à pessoa humana oriundos do uso de perfis falsos em sites de redes sociais

As transformações sociais advindas da popularização de Tecnologias da Informação e Comunicação (TICs) como a Internet são flagrantes. E a regulamentação destas novas práticas deve ser condizente com as peculiaridades das relações jurídicas desenvolvidas neste âmbito. Este trabalho visa analisar o problema da responsabilidade civil por danos à pessoa humana sofridos no âmbito dos sites de redes sociais pela criação e uso de perfis falsos. Assim, primeiramente, faz-se uma análise introdutória do fenômeno da Internet e das visões sobre as possíveis formas de regulamentação jurídica das condutas praticadas por meio da Rede. Posteriormente, adstringe-se à temática dos danos morais, buscando-se conceituá-los sob a ótica civil constitucional, como danos à pessoa humana. Finalmente, chega-se ao estudo dos perfis falsos, enfrentando-se a problemática dos danos morais sofridos por seu manejo. São desdobradas em três subproblemas: se são ou não capazes civilmente os usuários, chegando-se ao estudo dos perfis de menores de idade; se existem ou não as pessoas representadas nos perfis, subdividindo-se a análise nas hipóteses de perfis criados em roubo de identidade e perfis que representem criações intelectuais e; se estão ou não vivas as pessoas descritas nos perfis, trazendo-se à baila a hipótese dos perfis de pessoas falecidas.

Co-managerial potentials for Tanzania's Lake Victoria fisheries perspectives from two landing sites

Two landing sites were chosen in Tanzania for the 4-beaches survey. The former, Ihale, is a large one with an avarage of 120 boats and direct connections to the fish processing factories. The latter, Mwasonge, is one of the smallest landing sites in the Mwanza region with totally different characteristics. This book section aims to analyse the results from these two sites in the context of the co-management potentialities in Tanzania and more generally on the Lake Victoria region.

Heterometallic Complexes as Models of Enzymatic Active Sites

This dissertation describes studies on two multinucleating ligand architectures: the first scaffold was designed to support tricopper complexes, while the second platform was developed to support tri- and tetrametallic clusters.

In Chapter 2, the synthesis of yttrium (and lanthanide) complexes supported by a tripodal ligand framework designed to bind three copper centers in close proximity is described. Tricopper complexes were shown to react with dioxygen in a 1:1 [Cu₃]/O₂ stoichiometry to form intermediates in which the O–O bond was fully cleaved, as characterized via UV-Vis spectroscopy and determination of the reaction stoichiometry. Pre-arrangement of the three Cu centers was pivotal to cooperative O₂ activation, as mono-copper complexes reacted differently with dioxgyen. The reactivity of the observed intermediates was studied with various substrates (reductants, O-atom acceptors, H-atom donors, Brønsted acids) to determine their properties. In Chapter 3, the reactivity of the same yttrium-tricopper complex with nitric oxide was explored. Reductive coupling to form a trans-hyponitrite complex (characterized by X-ray crystallography) was observed via cooperative reactivity by an yttrium and a copper center on two distinct tetrametallic units. The hyponitrite complex was observed to release nitrous oxide upon treatment with a Brønsted acid, supporting its viability as an intermediate in nitric oxide reduction to nitrous oxide.

In Chapter 4, a different multinucleating ligand scaffold was employed to synthesize heterometallic triiron clusters containing one oxide and one hydroxide bridges. The effects of the redox-inactive, Lewis acidic heterometals on redox potential was studied by cyclic voltammetry, unveiling a linear correlation between redox potential and heterometal Lewis acidity. Further studies on these complexes showed that the Lewis acidity of the redox-inactive metals also affected the oxygen-atom transfer reactivity of these clusters. Comparisons of this reactivity with manganese systems, collaborative efforts to reassign the structures of related manganese oxo-hydroxo clusters, and synthetic attempts to access related dioxo clusters are also described.

In Appendix A, ongoing efforts to synthesize new clusters supported by the same multinucleating ligand platform are described. Studies of novel approaches towards ligand exchange in tetrametallic clusters and incorporation of new supporting and bridging ligand motifs in trinuclear complexes are presented.

La divulgación de información financiera en las páginas web de las empresas cotizadas

[ES] Es obligatorio para las empresas cotizadas que cuenten con una página web para atender el derecho de información de los accionistas, y donde difundir información relevante y obligatoria. Por ello, se creó la normativa necesaria para regular tanto la información mínima que las empresas debían aportar en sus webs, como los requisitos técnicos y jurídicos que dichas páginas debían poseer. No obstante, todas las empresas no presentan la misma información en sus webs, ya que muchas no se limitan a presentar solo la exigida por ley, y ésta puede ser más o menos útil para los usuarios. Por eso, es imprescindible poder conocer de alguna forma la transparencia que las empresas muestran a los usuarios en sus páginas web y en particular a los accionistas, si existe asimetría informativa o no, y saber cuál es el grado de calidad de la información presentada, así como la confianza que se puede tener en la misma.

Modelos de negocio innovadores en torno a la Web 2.0

El trabajo estudia la Web 2.0 y la repercusión que tiene a nivel empresarial. Idioma: Español.

Galveston Bay Migrates to the Web; DSpace Comes to Texas

This presentation describes how the Galveston Bay Bibliography and Galveston Bay Information Center projects transitioned from a paper world to electronic format. The mission of the Galveston Bay Information Center (GBIC) is to serve as a repository for information and a resource for research on Galveston Bay, its watershed, and the coastal region for all members of the Galveston Bay Community. GBIC was created in response to critical losses of data and information that were identified in the early stages of Galveston Bay National Estuary Program and includes a physical collection as well as a bibliography. With the loss of funding imminent, the administrators of GBIC recognized the need to preserve the collection and its digital bibliography in perpetuity. Enter Texas Digital Library. The University’s participation in TDL enabled GBIC to make the Galveston Bay Bibliography more robust and visible to the entire world.